Further Characterization of In Vitro Conditions Appropriate for GABA Determination in Human CSF: Impact of Acid Deproteinization and Freeze/Thaw

Thomas N. Ferraro, Bala V. Manyam, Theodore A. Hare

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21 Scopus citations

Abstract

Abstract: Recently established standardized protocols for collection, handling, and storage of CSF for measurement of γ‐aminobutyric acid (GABA) have proven valuable in the characterization of various CNS disorders. In response to two recent reports which may have an impact on certain widely used protocols, we have, using the confirmed ion‐exchange/fluorometric procedure, systematically evaluated the effects of deproteinization with various concentrations of sulfosalicylic acid (SSA) ranging from 0 to 10% (100 mg/ml), as well as the effects of freeze/ thaw (F/T) on CSF GABA levels. Results of F/T studies documented that levels are stable to freezing and thawing. Acid deproteinization studies revealed the presence of an equilibrium between strictly free GABA, demonstrable only in acid‐free CSF, and a very loosely bound form of GABA, fully demonstrable only in CSF deproteinized with concentrations of SSA above 1% (10 mg/ml). The relationship between GABA concentrations in undeproteinized and acid‐deproteinized CSF revealed a highly significant (p < .001) correlation, suggesting that alterations of central GABAergic activity would be reflected by either the level of strictly free GABA or free plus loosely bound GABA. This hypothesis was upheld in studies of patients with Parkinson's disease (PD) and Huntington's disease (HD), two neurologic disorders in which dysfunctions of the GABA system have been implicated. Results indicated that CSF GABA levels are significantly reduced in both PD and HD patients compared with neurologically normal controls, whether the measurement is of free GABA or free plus loosely bound GABA. Thus, we conclude that the level of strictly free GABA is stable to freezing and thawing and can only be accurately determined in nonacidified CSF; however, existing protocols employing deproteinization in 5% SSA yield data that provide an equally good reflection of central GABAergic transmission.

Original languageEnglish (US)
Pages (from-to)1057-1064
Number of pages8
JournalJournal of neurochemistry
Volume41
Issue number4
DOIs
StatePublished - Jan 1 1983
Externally publishedYes

All Science Journal Classification (ASJC) codes

  • Cellular and Molecular Neuroscience
  • Biochemistry

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